A water-soluble preparation for intravenous administration of isorhamnetin and its pharmacokinetics in rats.

Flavonoids are considered as health-protecting food constituents. The testing of their biological effects is however hampered by their low oral absorption and complex metabolism. In order to investigate the direct effect(s) of unmetabolized flavonoid, a preparation in a biologically friendly solvent for intravenous administration is needed. Isorhamnetin, a natural flavonoid and a human metabolite of the most frequently tested flavonoid quercetin, has very low water solubility (<3.5 µg/mL). The aim of this study was to improve its solubility to enable intravenous administration and to test its pharmacokinetics in an animal model. By using polyvinylpyrrolidone (PVP10) and benzalkonium chloride, we were able to improve the solubility approximately 600 times to 2.1 mg/mL. This solution was then administered intravenously at a dose of 0.5 mg/kg of isorhamnetin to rats and its pharmacokinetics was analyzed. The pharmacokinetics of isorhamnetin corresponded to two compartmental model with a rapid initial distribution phase (t1/2α: 5.7 ± 4.3 min) and a slower elimination phase (t1/2ß: 61 ± 47.5 min). Two sulfate metabolites were also identified. PVP10 and benzalkonium did not modify the properties of isorhamnetin (iron chelation and reduction, and cell penetration) substantially. In conclusion, the novel preparation reported in this study is suitable for future testing of isorhamnetin effects under in vivo conditions.

Mucoadhesive phenylboronic acid-grafted carboxymethyl cellulose hydrogels containing glutathione for treatment of corneal epithelial cells exposed to benzalkonium chloride.

Benzalkonium chloride (BAK) is the most commonly-used preservative in topical ophthalmic medications that may cause ocular surface inflammation associated with oxidative stress and dry eye syndrome. Glutathione (GSH) is an antioxidant in human tears and able to decrease the proinflammatory cytokine release from cells and reactive oxygen species (ROS) formation. Carboxymethyl cellulose (CMC), a hydrophilic polymer, is one of most commonly used artificial tears and can promote the corneal epithelial cell adhesion, migration and re-epithelialization. However, most of commercial artificial tears provide only temporary relief of irritation symptoms and show the short-term treatment effects. In the study, 3-aminophenylboronic acid was grafted to CMC for increase of mucoadhesive properties that might increase the precorneal retention time and maintain the effective therapeutic concentration on the ocular surface. CMC was modified with different degree of substitution (DS) and characterized by Fourier transform infrared spectroscopy and nuclear magnetic resonance spectroscopy. Phenylboronic acid (PBA)-grafted CMC hydrogels have interconnected porous structure and shear thinning behavior. Modification of CMC with high DS (H-PBA-CMC) shows the strong bioadhesive force. The optimal concentration of GSH to treat corneal epithelial cells (CECs) was evaluated by cell viability assay. H-PBA-CMC hydrogels could sustained release GSH and decrease the ROS level. H-PBA-CMC hydrogels containing GSH shows the therapeutic effects in BAK-damaged CECs via improvement of inflammation, apoptosis and cell viability. After topical administration of developed hydrogels, there was no ocular irritation in rabbits. These results suggested that PBA-grafted CMC hydrogels containing GSH might have potential applications for treatment of dry eye disease.

Generic benzalkonium chloride-preserved travoprost eye drops are not identical to the branded polyquarternium-1-preserved travoprost eye drop: Effect on cultured human conjunctival goblet cells and their physicochemical properties.

PURPOSE: To investigate the effect of polyquaternium-1 (PQ)-preserved and benzalkonium chloride (BAK)-preserved travoprost eye drops on viability of primary human conjunctival goblet cell (GC) cultures and on secretion of mucin and cytokines. Furthermore, to evaluate the physicochemical properties of the branded travoprost eye drop Travatan® and available generics. METHODS: The effect of travoprost eye drops was evaluated on GC cultures. Cell viability was assessed through lactate dehydrogenase (LDH) and tetrazolium dye (MTT) colorimetric assays. Mucin secretion was evaluated by immunohistochemical staining. Secretion of interleukin (IL)-6 and IL-8 was measured using BD Cytometric Bead Arrays. pH, viscosity, droplet mass, osmolality and surface tension were measured for all included eye drops. RESULTS: In the LDH assay, BAK travoprost caused significant GC loss after 2 hrs of incubation compared to the control. PQ travoprost caused no GC loss at any time point. Both PQ- and BAK travoprost caused secretion of mucin to the cytoplasma. No difference in IL-6 and IL-8 secretion was identified compared to controls. The pH values for the generics were lower (pH 6.0) than the pH value for Travatan (pH 6.7; p < 0.0001). The viscosity was lowest for Travatan, while the mean droplet mass was higher for Travatan (35 mg) than the generics (28-30 mg; p ≤ 0.0318). The osmolality and surface tension did not differ between the eye drops investigated. CONCLUSION: BAK travoprost caused GC loss, indicating that PQ preservation may be preferable in treatment of glaucoma. Furthermore, physicochemical properties of branded and generic travoprost eye drops can not be assumed to be identical.

Lipophilic Arginine Esters: The Gateway to Preservatives without Side Effects.

This study hypothesized that long carbon chain cationic arginine (Arg) esters can be considered as toxicologically harmless preservatives. Arg-esters with C18 and C24 carbon chains, namely, arginine-oleate (Arg-OL) and arginine-decyltetradecanoate (Arg-DT), were synthesized. Structures were confirmed by FT-IR, 1H NMR, and mass spectroscopy. Both Arg-esters were tested regarding hydrophobicity in terms of log Poctanol/water, critical micelle concentration (CMC), biodegradability, cytotoxicity, hemolysis, and antimicrobial activity against Escherichiacoli (E. coli), Staphylococcusaureus (S. aureus), Bacillussubtilis (B. subtilis), and Enterococcusfaecalis (E. faecalis). Log Poctanol/water of arginine was raised from -1.9 to 0.3 and 0.6 due to the attachment of C18 and C24 carbon chains, respectively. The critical micelle concentration of Arg-OL and Arg-DT was 0.52 and 0.013 mM, respectively. Both Arg-esters were biodegradable by porcine pancreatic lipase. In comparison to the well-established antimicrobials, benzalkonium chloride (BAC) and cetrimide, Arg-esters showed significantly less cytotoxic and hemolytic activity. Both esters exhibited pronounced antimicrobial properties against Gram-positive and Gram-negative bacteria comparable to that of BAC and cetrimide. The minimum inhibitory concentration (MIC) of Arg-esters was <50 µg mL-1 against all tested microbes. Overall, results showed a high potential of Arg-esters with long carbon chains as toxicologically harmless novel preservatives.

Benzalkonium chloride and cetylpyridinium chloride induce apoptosis in human lung epithelial cells and alter surface activity of pulmonary surfactant monolayers.

Quaternary ammonium compounds (e.g., benzalkonium chloride (BAC) and cetylpyridinium chloride (CPC)) constitute a group of cationic surfactants are widely used for personal hygiene and medical care despite the potential pulmonary toxicity. To examine whether BAC and CPC aerosols deposited in the alveolar region alter pulmonary function, we studied the effects on pulmonary surfactant using two-step in vitro models; cytotoxicity using A549 alveolar epithelial cell and changes in surface activity of the pulmonary surfactant monolayer using both Surfacten® and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). Cell viability was decreased with BAC and CPC dose-dependently. A comparison of cytotoxicity among BAC homologues with different length of alkyl chain showed that C16-BAC, which has the longest alkyl chain, was more cytotoxic than C12- or C14-BAC. Caspase-3/7 activity and cleaved form of caspase-3 and PARP were increased in BAC- and CPC-exposed cells. The elevated caspase-3/7 activity and their cleaved active forms were abolished by caspase-3-inhibitor. Furthermore, we examined the features of the surface pressure/trough area (π-A) isotherm by the Langmuir-Wilhelmy method and atomic force microscopy (AFM) images of lipid monolayers on a subphase containing BAC, CPC, or pyridinium chloride (PC, as a control). The π-A isotherms showed that addition of BAC or CPC yielded dose-dependent increases in surface pressure without compression, indicating that BAC and CPC expand the isotherm to larger areas at lower pressure. The collapse pressure diminished with increasing concentration of CPC. Topographic images indicated that BAC and CPC resulted in smaller condensed lipid domains compared to the control. Conversely, PC without hydrocarbon tail group, showed no cytotoxicity and did not change the isotherms and AFM images. These results indicate that BAC and CPC cause cell death via caspase-3-dependent apoptotic pathway in A549 cells and alter the alveolar surfactant activity. These effects can be attributed to the long alkyl chain of BAC and CPC.

One-step hydrophobization of tannic acid for antibacterial coating on catheters to prevent catheter-associated infections.

Catheter-associated infections (CAIs) caused by bacterial colonization are significant problems in clinics. Thus, effective antibacterial coatings for biomedical catheters to prevent bacterial infections are urgently needed. Ideal coatings should include the advantage of potent antibacterial properties and being easily and economically modified on the catheter surface. Due to their advantages of adhesive capability on various substrates, an increasing number of coatings based on plant polyphenols have been developed. However, the hydrophilicity of plant polyphenols limits their utilization in coatings. Herein, hydrophobic tannic acid (TA) was synthesized via the one-step electrostatic assembly of TA and benzalkonium chloride (BAC) with the green solvent water as the medium. The as-prepared hydrophobic TA (TBA) facilely formed a stable and colorless coating on the luminal and outer surface of biomedical catheters with broad-spectrum antibacterial activity and biocompatiblity. It was demonstrated that the TBA-coated surfaces displayed excellent bactericidal activity toward Gram-positive Staphylococcus aureus (S. aureus) and Gram-negative Escherichia coli (E. coli), and more than 99% of the above bacteria were killed by the TBA-coated films. The test of the coated catheters in vitro also showed the excellent antibacterial activity of both the outer and luminal surfaces of the catheter. Moreover, in an in vivo mouse model, the coated catheters relatively prevented bacterial colonization compared to the uncoated catheters. Meantime, no significant cytotoxicity and host response for Cell Counting Kit-8 (CCK-8) and tissue compatibility in vivo were observed, indicating the better biocompatibility of the TBA coating. This preparation method overcomes the limitation of the traditional hydrophilic tannic acid as a coating and provides a new method for preventing medical indwelling device-associated infections.

The preparation and morphology control of heparin-based pH sensitive polyion complexes and their application as drug carriers.

Heparin as negative polysaccharide is a universal building block to form polyion complex with different cationic counterparts. In this paper, three different cations, including chitosan, benzyldodecyldimethyl ammonium bromide and doxorubicin hydrochloride, were used to prepare heparin-based polyion complexes (HPICs). Their morphologies could be tuned by heparin content in HPIC, and they also showed pH-sensitive decomposition. Doxorubicin was further encapsulated into micelle and vesicle carrier made from heparin-benzyldodecyl dimethyl ammonium bromide PIC, whereas heparin-doxorubicin PIC could be directly used as drug carrier. In vitro drug release proved the drug carriers exhibit obvious pH sensitive release behaviour. Cytotoxicity indicated the drug carrier possessed significant cytotoxicity to tumor cells. The cell uptake observed by CLSM showed the carrier was able to deliver antitumor drug into tumor cell's nucleus. Consequently, these results showed the promising potential of HPIC in drug carrier application.

Influência de inibidores de metaloproteinases na degradação da camada híbrida / Influence of metalloproteinase inhibitors on hybrid layer degradation

Objetivo: realizar uma revisão de literatura acerca da eficácia de utilização da clorexidina (CHX) e de outros tipos de inibidores de metaloproteinases (MMPs) na resistência de união da camada híbrida. Métodos: a busca bibliográfica foi realizada na base de dados PubMed, nos meses de novembro e dezembro de 2018. A pesquisa ocorreu em três fases, com os descritores previamente selecionados. Foram incluídas publicações dos últimos 10 anos no formato de pesquisas científicas realizadas in vitro ou in vivo. Após análise, obedecendo aos critérios de inclusão e exclusão, foram incluídos sete estudos na presente revisão. Resultados/Revisão de literatura: na interface adesiva, os estudos mostram que as MMPs são ativadas durante a etapa de ataque ácido realizada nos protocolos de aplicação de sistemas adesivos, podendo ser ativada tanto por procedimentos adesivos com condicionamento ácido prévio como por sistemas adesivos autocondicionantes. Além da CHX, outras substâncias foram pesquisadas e se mostraram eficazes na inibição de MMPs. Considerações finais: por meio da inibição da atividade das MMPs, é possível obter uma maior durabilidade da interface adesiva e uma menor degradação hidrolítica do colágeno presente na camada híbrida. (AU)

Objective: to perform a literature review on the efficacy of chlorhexidine (CHX) and other types of metalloproteinase inhibitors (MMPs) on hybrid layer bond strength. Methods: the bibliographic search was performed in PubMed, in the months of november and december of 2018. The research was carried out in three phases with the previously selected descriptors. Publications have been included in the last 10 years in the form of scientific research conducted in vitro or in vivo. After analysis, following the inclusion and exclusion criteria, 7 studies were included in the present review. Results / Literature review: in the adhesive interface, the studies show that the MMPs are activated during the acid attack stage carried out in the application protocols of adhesive systems, and can be activated either by adhesive procedures with prior acid conditioning or self-etching adhesive systems. In addition to CHX, other substances were investigated and shown to be effective in inhibiting MMPs. Final considerations: through the inhibition of the MMPs activity it is possible to obtain a greater durability of the adhesive interface and lower hydrolytic degradation of the collagen present in the hybrid layer. (AU)

Influence of surfactants addition on the properties of calcium hypochlorite solutions

Abstract Objectives The aim of this study was to evaluate the influence of surfactants 0.2% or 0.1% cetrimide (Cet) or 0.008% benzalkonium chloride (BAK) on 2.5% calcium hypochlorite (Ca(OCl)2), and compare to sodium hypochlorite (NaOCl), regarding the properties of pH, free chlorine content, surface tension, contact angle, pulp dissolution and antimicrobial activity. Material and Methods The pH and free chlorine content were evaluated by digital pHmeter and by titration, respectively. Surface tension was measured by the platinum ring technique with a Du Noüy tensiometer. The solution's contact angle in human dentin surfaces was checked by Drop Shape Analyzer software. Bovine pulps were used for pulp dissolution analysis and the dissolving capacity was expressed by percent weight loss. Antimicrobial activity over Enterococcus faecalis was evaluated by the agar diffusion method. Results Surfactants addition to Ca(OCl)2 and NaOCl did not alter the pH, free chlorine content and pulp dissolution properties. Ca(OCl)2 had the highest surface tension among all tested solutions. When surfactants were added to Ca(OCl)2 and NaOCl, there was a significant reduction of surface tension and contact angle values. The addition of 0.2% or 0.1% Cet enhanced antimicrobial activity of both Ca(OCl)2 and NaOCl. Conclusion Surfactant addition to 2.5% Ca(OCl)2 has shown acceptable outcomes for pH, free chlorine content, surface tension, contact angle, pulp dissolution and antimicrobial activity. Furthermore, the addition of 0.2% Cet showed better results for all tested properties.

Mucoadhesive self-emulsifying delivery systems for ocular administration of econazole.

AIM: Development of mucoadhesive self-emulsifying drug delivery systems (SEDDS) providing a prolonged ocular residence time for poorly soluble active pharmaceutical ingredient. METHODS: l-Cysteine was covalently linked to 6-mercaptonicotinamide. The obtained ligand, Cysteine-6-mercaptonicotinamide (Cys-6-MNA) was attached to Eudragit® L100-55 via a carbodiimide mediated amide bond formation. The resulting entirely S-protected thiolated Eudragit® L100-55 was characterized regarding the degree of modification as well as stability toward oxidation in the presence of strong oxidizing agent (H2O2). The S-protected thiolated Eudragit® L100-55 was incorporated into SEDDS via hydrophobic ion pairing with benzalkonium chloride (BAK) in a concentration of 2% (m/m). S-protected thiolated Eudragit® L100-55-BAK ion pair SEDDS (S-protected thiolated EU-BAK SEDDS) were characterized regarding their physicochemical and mucoadhesive properties. Econazole nitrate (EN) was incorporated into SEDDS in concentration of 1% (m/m) and in vitro drug release was assessed. Furthermore, toxicity study was performed on procine corneas via resazurin assay. RESULTS: The entirely S-protected thiolated Eudragit® L100-55 exhibited 282 ±â€¯78.25 µmol of MNA per gram of polymer. Ellman's test confirmed no free thiol groups and stability study showed no significant increase in dynamic viscosity overtime. The droplet size of developed SEDDS in simulated lacrimal fluid was below 100 nm with polydispersity index below 0.3. S-protected thiolated EU-BAK SEDDS exhibited 2.5-fold higher mucoadhesive properties than blank SEDDS on ocular mucosa. S-protected thiolated EU-BAK SEDDS showed sustained EN release over period of 8 h and no pronounced corneal toxicity in 0.5% (m/v) concentration. CONCLUSION: Accordingly, these mucoadhesive SEDDS can be considered as promising ocular delivery system for EN.

Gold nanoparticles capped with benzalkonium chloride and poly (ethylene imine) for enhanced loading and skin permeability of 5-fluorouracil.

OBJECTIVE: To enhance 5-fluorouracil (5-FU) permeability through the skin by loading onto gold nanoparticles (GNPs) capped with two cationic ligands, benzalkonium chloride (BC) or poly (ethylene imine) (PEI). Whereas 5-FU has excellent efficacy against many cancers, its poor permeability through biological membranes and several adverse effects limit its clinical benefits. BC and PEI were selected to stabilize GNPs and to load 5-FU through ionic interactions. METHODS: 5-FU/BC-GNPs and 5-FU/PEI-GNPs were prepared at different 5-FU/ligand molar ratios and different pH values and were evaluated using different techniques. GNPs stability was tested as a function of salt concentration and storage time. 5-FU release from BC- and PEI-GNPs was evaluated as a function of solution pH. Ex vivo permeability studies of different 5-FU preparations were carried out using mice skin. RESULTS: 5-FU-loaded GNPs size and surface charge were dependent on the 5-FU/ligand molar ratios. 5-FU entrapment efficiency and loading capacity were dependent on the used ligand, 5-FU/ligand molar ratio and solution pH. Maximum drug entrapment efficiency of 59.0 ± 1.7% and 46.0 ± 1.1% were obtained for 5-FU/BC-GNPs and 5-FU/PEI-GNPs, respectively. 5-FU-loaded GNPs had good stability against salinity and after storage for 4 months at room temperature and at 4 °C. In vitro 5-FU release was pH- and ligand-dependent where slower release was observed at higher pH and for 5-FU/BC-GNPs. 5-FU permeability through mice skin was significantly higher for drug-loaded GNPs compared with drug-ligand complex or drug aqueous solution. CONCLUSION: Based on these results, BC- and PEI-GNPs might find applications as effective topical delivery systems of 5-FU.

Validation of the AccuPoint Advanced ATP Hygiene Monitoring System for Sanitation Monitoring Through Detection of ATP from Stainless Steel Surfaces.

The AccuPoint Advanced ATP Hygeine Monitoring System was validated by an AOAC International Performance Tested MethodSM on the detection of ATP from stainless steel surfaces. Neogen Corp.'s system is a lightweight, hand-held diagnostic tool used to validate and verify a hygiene program's effectiveness by detecting organic residues remaining on surfaces and in liquids after cleaning. The system is composed of three primary components: an electronic luminometer, fully self-contained single-use samplers, and software. The system is designed to detect adenosine triphosphate (ATP) at set thresholds and to report the measurement in relative light units (RLU). These thresholds are established by a facility to reflect effective cleaning practices. The instrument compares the measured level of ATP with the established threshold and reports the results as pass, marginal, or fail. A linear dose-response in RLU was observed with pure analyte. In the matrix and microbial studies, detection levels varied depending on the matrix and microorganism tested. Independent laboratory trials confirmed pure analyte and matrix observations. Specificity testing of similar, yet different, compounds resulted in 0 RLU for all except 2'-deoxyadenosine 5'-triphosphate sodium salt, which showed markedly reduced reactivity when compared with ATP. Also, interference by these compounds was negligible. When disinfectant residues were evaluated for their effect on the test, cleaners increased RLU output to varying degrees. Stability testing showed consistent results between three independently manufactured lots and stable results through the 9 month shelf-life. Additionally, when three readers were compared using electronic light-emitting diodes as the light source, instrument variability was low (<3%). Robustness testing results provided evidence that temperature affects test performance more than shaking time, and sampler performance improves as the temperature increases to room temperature. These results provided evidence that the AccuPoint Advanced ATP Hygiene Monitoring System produces consistent and reliable data for the evaluation of sanitation program effectiveness on stainless steel surfaces in food processing and food service facilities.

Headspace gas chromatography based methodology for the analysis of aromatic substituted quaternary ammonium salts.

The analysis of quaternary ammonium salts (QAS) using GC is often performed by "in injector" pyrolysis to create volatile degradation products for quantification purposes. Besides the risk of severe system contamination, the application of this approach on aqueous samples is problematic. In this work, the sample is treated in a vial with 2,2-dimethoxypropane (DMP) under acidic catalysis. In addition to the removal of water and sample enrichment, the QAS are decomposed. As HS transfers only volatile compounds to the GC system, contamination is avoided. It was found that depending on the presence of benzyl, phenyl or methyl groups on the quaternary nitrogen; benzyl chloride, N,N-dimethylaniline or chloromethane are formed respectively in the sealed vial. All these can be used as an analytical target. A calibration curve for benzyl chloride could be derived from the pure compound. Chloromethane was generated from pure benzyldimethyldecylammonium chloride (BEDIDE), a pure QAS with benzyl and methyl groups, to construct a secondary calibration curve using a back analysis approach. It has been proven that by quantifying the formed analytical targets, the mass balance for the QAS under investigation was close to 100%. The presented procedure allows the quantification of any aromatic substituted QAS without the need for a matching reference, which is a major advantage over existing CE and LC methods The proposed methodology was validated for mouth sprays containing benzethonium chloride (BZTCl) or benzoxonium chloride (BZOCl) and for denatonium benzoate (DB) in ethylene glycol (EG) based cooling liquids. Results showed that the approach provided excellent linearity (R2≥0.999) and limits of detection around 0.01µg/vial for benzyl chloride. It was found that the reaction product of DMP and glycerol which was also present in the mouthspray and some cooling liquids, caused chromatographic interference with benzyl chloride. Treating those samples in the vial with N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) after the enrichment step removes the interference and leaves a possible pathway for the simultaneous determination of glycerol in those samples.

A Water-Soluble Cyclotriveratrylene-Based Supra-amphiphile: Synthesis, pH-Responsive Self-Assembly in Water, and Its Application in Controlled Drug Release.

A new water-soluble cyclotriveratrylene (WCTV) was designed and synthesized, and benzyldimethyldodecylammonium chloride (G) was chosen as the guest molecule to construct a supra-amphiphile by the host-guest interaction between WCTV and G in water, which is pH responsive. The supra-amphiphiles self-assembled into vesicles in water. When the pH of the solution was below 7.0, the supra-amphiphile disassociated, and the vesicles collapsed. Then, the pH-responsive self-assembly system was utilized for controlled drug release.

Identification of Environmental Quaternary Ammonium Compounds as Direct Inhibitors of Cholesterol Biosynthesis.

In this study, we aim to identify environmental molecules that can inhibit cholesterol biosynthesis, potentially leading to the same biochemical defects as observed in cholesterol biosynthesis disorders, which are often characterized by congenital malformations and developmental delay. Using the Distributed Structure-Searchable Toxicity (DSSTox) Database Network developed by EPA, we first carried out in silico screening of environmental molecules that display structures similar to AY9944, a known potent inhibitor of 3ß-hydroxysterol-Δ(7)-reductase (DHCR7)-the last step of cholesterol biosynthesis. Molecules that display high similarity to AY9944 were subjected to test in mouse and human neuroblastoma cells for their effectiveness in inhibiting cholesterol biosynthesis by analyzing cholesterol and its precursor using gas chromatography-mass spectrometry. We found that a common disinfectant mixture, benzalkonium chlorides (BACs), exhibits high potency in inhibiting DHCR7, as suggested by greatly elevated levels of the cholesterol precursor, 7-dehydrocholesterol (7-DHC). Subsequent structure-activity studies suggested that the potency of BACs as Dhcr7 inhibitors decrease with the length of their hydrocarbon chain: C10 > C12 ≫ C14 > C16. Real-time qPCR analysis revealed upregulation of the genes related to cholesterol biosynthesis and downregulation of the genes related to cholesterol efflux, suggesting a feedback response to the inhibition. Furthermore, an oxidative metabolite of 7-DHC that was previously identified as a biomarker in vivo was also found in cells exposed to BACs by liquid chromatography-mass spectrometry. Our findings suggest that certain environmental molecules could potently inhibit cholesterol biosynthesis, which could be a new link between environment and developmental disorders.

Towards an antimicrobial 'microglove'.

A large proportion of hospital-related infections are acquired and spread due to the direct contacts between patients and healthcare workers. Accordingly, proper infection prevention measures, and especially hand hygiene, are key to limit the spread of infections in nosocomial settings. However, healthcare workers frequently experience difficulties in complying strictly to hand disinfection protocols. This study was therefore aimed at the development of a hand rub with antimicrobial activity that forms a protective film on the hand, a so-called microglove, in order to enhance hand hygiene. For this purpose, various co-polymer formulations consisting of different ratios of Polyvinylpyrrolidone (PVP) and a branched C20 derivatized maleate (M20) in combination with the known biocide benzalkonium chloride (BKC) were tested for their combined film-forming and antimicrobial activities. The results of a series of novel contamination and transmission assays show that a formulation of 80% PVP and 20% M20 co-polymer with 0.9% BKC fulfils the elementary requirements for an antimicrobial microglove.

Development of a cyclodextrin-based aqueous cyclosporin A eye drop formulations.

Cyclosporin A (CyA) is a lipophilic, cyclic polypeptide drug with anti-inflammatory properties. It is used in topical treatment of dry eyes and is now commercially available in oil based surfactant containing eye drops. Surfactants can irritate the eye surface causing burning, itching and irritation of the conjunctiva, and oil-based drops can result in blurred vision. Thus, the aim of this study was to develop surfactant free aqueous 0.05% (w/v) CyA eye drops where the drug is present in an aqueous vehicle containing CyA/cyclodextrin (CyA/CD) nanoparticles. The effects of the natural α-, ß- and γ-cyclodextrins (αCD, ßCD and γCD), as well as of the water soluble hydroxypropyl derivatives of γCD and αCD (HPγCD, HPαCD) and randomly methylated ßCD (RMßCD), were determined in pure water. αCD had the best solubilizing effect increasing the solubility of CyA above 0.05% upon addition of only 5% (w/v) αCD. γCD did not have as good solubilizing effect but was tested further due to its superior ability to form nanoparticles and its favorable toxicological profile. Seven eye drop formulations were prepared and tested. All contained 0.05% (w/v) CyA in addition to polyvinyl alcohol, benzalkonium chloride, disodium edetate and various amounts of CD (αCD, γCD and mixtures thereof). When the formulation contained only αCD most of the drug was dissolved but some small aggregates were formed with hydrodynamic diameter of about 6 and 155 nm. When the formulation contained only γCD negligible CyA/CD complexation occurred with most of the drug present as solid CyA particles. When the formulation contained a mixture of αCD and γCD, where αCD concentration was at least 3% (w/v), the entire drug content was dissolved in the media under formation of relatively large (100-2000 nm) CyA/CD nanoparticles. αCD solubilized the drug while γCD enhanced nanoparticle formation. The effect of polyvinyl alcohol, benzalkonium chloride and disodium edetate on the nanoparticle formation was also investigated and shown to have positive effect on the aggregate formation.

Optimising concentrations of antimicrobial agents in pharmaceutical preparations: Case of an oral solution of glycerol and an ophthalmic solution containing cysteamine.

INTRODUCTION: In the context of current distrust of antimicrobial preservatives, the quantities of these substances in two pharmaceutical formulas were studied: an ophthalmic solution of cysteamine preserved benzalkonium chloride at 1mg/5mL and Glycerotone(®) preserved with sorbic acid at 0.1g/100g. The purpose of this work was to verify that a reduction of the quantities of preservative continues to fulfil the requirements for antimicrobial preservation. MATERIAL AND METHODS: The Test of efficacy of antimicrobial preservation, section 5.1.3 of the 8th edition of the European Pharmacopoeia, was carried out on each formulation prepared with decreasing quantities of preservative. RESULTS: The results show that formulations whose preservative concentration was reduced by a factor of four remained compliant with standards. It is to be noted that in formulas without preservative, fungal growth was observed in both the solution of Glycerotone(®) and the ophthalmic solution containing cysteamine. DISCUSSION: Although there is no question that an antimicrobial preservative is necessary, the quantity of preservative can be reduced without deteriorating the quality of the pharmaceutical product but the minimal effective concentration remains to be determined. CONCLUSION: The formulations of many pharmaceutical products should therefore be examined in order to limit the quantities of preservative while continuing to guarantee patient's safety.

Preventive and therapeutic effects of SkQ1-containing Visomitin eye drops against light-induced retinal degeneration.

The human retina is constantly affected by light of varying intensity, this being especially true for photoreceptor cells and retinal pigment epithelium. Traditionally, photoinduced damages of the retina are induced by visible light of high intensity in albino rats using the LIRD (light-induced retinal degeneration) model. This model allows study of pathological processes in the retina and the search for retinoprotectors preventing retinal photodamage. In addition, the etiology and mechanisms of retina damage in the LIRD model have much in common with the mechanisms of the development of age-related retinal disorders, in particular, with age-related macular degeneration (AMD). We have studied preventive and therapeutic effects of Visomitin eye drops (based on the mitochondria-targeted antioxidant SkQ1) on albino rat retinas damaged by bright light. In the first series of experiments, rats receiving Visomitin for two weeks prior to illumination demonstrated significantly less expressed atrophic and degenerative changes in the retina compared to animals receiving similar drops with no SkQ1. In the second series, the illuminated rats were treated for two weeks with Visomitin or similar drops without SkQ1. The damaged retinas of the experimental animals were repaired much more effectively than those of the control animals. Therefore, we conclude that Visomitin SkQ1-containing eye drops have pronounced preventive and therapeutic effects on the photodamaged retina and might be recommended as a photoprotector and a pharmaceutical preparation for the treatment of AMD in combination with conventional medicines.

In vitro effects of preserved and unpreserved anti-allergic drugs on human corneal epithelial cells.

PURPOSE: Treatment with topical eye drops for long-standing ocular diseases like allergy can induce detrimental side effects. The purpose of this study was to investigate in vitro cytotoxicity of commercially preserved and unpreserved anti-allergic eye drops on the viability and barrier function of monolayer and stratified human corneal-limbal epithelial cells. METHODS: Cells were treated with unpreserved ketotifen solution, benzalkonium chloride (BAC)-containing anti-allergic drugs (ketotifen, olopatadine, levocabastine) as well as BAC alone. 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was used to determine cell viability. Effects of compounds on barrier function were analyzed measuring transepithelial electrical resistance (TEER) to determine paracellular permeability and rose bengal assays to evaluate transcellular barrier formation. RESULTS: The BAC-preserved anti-allergic formulations and BAC alone significantly reduced cell viability, monolayer cultures being more sensitive to damage by these solutions. Unpreserved ketotifen induced the least diminution in cell viability. The extent of decrease of cell viability was clearly dependent of BAC presence, but it was also affected by the different types of drugs when the concentration of BAC was low and the short time of exposure. Treatment with BAC-containing anti-allergic drugs and BAC alone resulted in increased paracellular permeability and loss of transcellular barrier function as indicated by TEER measurement and rose bengal assays. CONCLUSIONS: The presence of the preservative BAC in anti-allergic eye drop formulations contributes importantly to the cytotoxic effects induced by these compounds. Stratified cell cultures seem to be a more relevant model for toxicity evaluation induced on the ocular surface epithelia than monolayer cultures.